【摘要】 目的观察神经营养因子与神经干细胞(NSCs)联合移植对缺氧缺血性脑损伤(HIBD)大鼠的治疗效果。方法取新生24h的Wistar大鼠海马NSCs进行培养、鉴定。选7日龄Wistar大鼠制备HIBD动物模型,7d后行损伤侧侧脑室移植。将实验大鼠随机分为9组:正常组、模型组、磷酸盐缓冲液移植组、NSCs移植组、BDNF组、BDNF+NSCs移植组、NT-3组、NT-3+NSCs移植组、BDNF+NT-3+NSCs移植组。移植4周后进行功能实验,取脑组织进行免疫组化及免疫荧光检查。结果从新生大鼠海马可成功培养出NSCs,并表达NSCs的标志物神经巢蛋白(neuroepi-thelial stem cell protein,Nestin),NSCs可分化为神经元及星形胶质细胞。Y迷宫实验:NSCs移植组达到学会的次数(163±11.60)n,正确记忆次数(5.00±1.13)n;BDNF组(150.00±8.94,5.17±1.47)n;BDNF+NSCs移植组(111.25±13.56,6.23±1.60)n;NT-3组(153.56±10.35,5.07±1.03)n;NT-3+NSCs移植组(117.27±11.4,6.30±1.1)n;BDNF+NT-3+NSCs移植组(110±11.55,6.30±1.1)n。悬吊实验:NSC移植组平均(30.10±11.8)s;BDNF组(36.25±10.98)s;BDNF+NSCs移植组(43.6±10.56)s;NT-3组(34.95±10.15)s;NT-3+NSCs移植组(40.64±10.6)s;BDNF+NT-3+NSCs移植组(42.20±6.25)s。斜坡试验:NSCs移植组平均(20.3±8.25)s;BDNF组(14.33±2.88)s;BDNF+NSC移植组(11.17±2.48)s;NT-3组(15.26±2.98)s;NT-3+NSC移植组(12.8±3.65)s;BDNF+NT-3+NSCs移植组(12.9±5.18)s。说明各联合移植组大鼠学习记忆能力及神经功能与NSCs移植组比较,明显改善(P<0.05),而NSCs组大鼠学习记忆能力及神经功能与HIBD组比较,差异无统计学意义(P>0.05)。各联合移植组损伤侧皮层、海马存活的NSCs数量(BDNF+NSC移植组9.31±0.71个,10.10±1.65个;NT-3+NSCs移植组6.18±1.83个,8.18±3.06个;BDNF+NT-3+NSCs移植组为9.58±1.61个,11.45±1.36个)与NSCs组(3.33±0.24个,4.22±0.33个)比较明显增多(P<0.05),且NSCs分化为神经元的比例明显增多,与NSCs移植组比较,有统计学意义(P<0.05),但双因子联合移植组与其单因子移植组比较,差异无统计学意义(P>0.05)。结论BDNF、NT-3与NSCs联合移植是治疗HIBD的有效方法。更多还原

【Abstract】 Objective To investigate the effects of brain-derived neurotrophic factor(BDNF), Neurotrophin-3 (NT-3) with neural stem cells (NSCs) transplantation in treatment of hypoxic-ischemic brain damage (HIBD)in newborn rats. Methods NSCs were separated from hippocampus of neonatal Wistar rat’s of 24 h age. Seven-days rats were sustained HIBD. Seven days later, animals received transplantation in injured side cerebral ventricle. They were randomly divided into normal group, HIBD group, PBS group, NSCs group, BDNF group, BDNF+NSCs group, NT-3 group, NT-3+NSCs group and BDNF+NT-3+NSCs group. Four weeks after transplantation, ability tests and histological exam were taken. Results The hippocampus NSCs of newborn rat were well cultured and they could express nestin, which was the marker of NSCs. NSCs could differentiate into NSE and GFAP. Y-maze test: attain grasping time (s) and correct memory time (s) in NSCs were (163±11.60, 5.00±1.13)n; BDNF group were (150.00±8.94, 5.17±1.47)n; BDNF+NSCs group were (111.25±13.56, 6.23±1.60)n; NT-3 group were (153.56±10.35, 5.07±1.03)n; NT-3+NSCs group were (117.27±11.4, 6.30±1.1)n; BDNF+NT-3+NSCs group were (110±11.55, 6.30±1.1)n; Hanging test duration(s): NSCs group was (30.10±11.8)s; BDNF group was (36.25±10.98)s; BDNF+NSCs group was (43.6±10.56)s; NT-3 group was (34.95±10.15)s; NT-3+NSCs group was (40.64±10.6)s; BDNF+NT-3+NSCs group was (42.20±6.25)s. Inclined plane test also showed better results in combined group. The learning and memory ability and neuron function of the associated transplantation groups were better improved than the NSCs group(P<0.05). But there was no difference between the NSCs group and model group (P>0.05).Compared with NSCs group (3.33±0.24, 4.22±0.33), both the number of survival NSCs in combined transplantation groups (BDNF+NSCs group 9.31±0.71, 10.10±1.65.NT-3+NSCs group 6.18±1.83, 8.18±3.06.BDNF+NT-3+NSCs group 9.58±1.61, 11.45±1.36) and the percentage of NSCs differentiating into NSE increased obviously (P<0.05). There was no significant difference between the dual factors transplantation group and the haplo-fator transplantation groups(P>0.05). Conclusions The combined transplantation of BDNF, NT-3 and NSCs is an effective method to treat HIBD.更多还原